Use of the phytocannabinoid cannabidivarin (cbdv) in the treatment of epilepsy

ABSTRACT

This invention relates to the use of the phytocannabinoid cannabidivarin (CBDV) and combinations of the phytocannabinoid CBDV with tetrahydrocannabivarin (THCV) and cannabidiol (CBD) in the treatment of epilepsy. The invention further relates to the use of the phytocannabinoid CBDV in combination with standard anti-epileptic drugs (SAEDs). Preferably the SAED is one of ethosuximide, valproate or phenobarbital.

FIELD OF THE INVENTION

This invention relates to the use of the phytocannabinoid cannabidivarin(CBDV) and combinations of the phytocannabinoid CBDV withtetrahydrocannabivarin (THCV) and cannabidiol (CBD) in the treatment ofepilepsy. The invention further relates to the use of thephytocannabinoid CBDV in combination with standard anti-epileptic drugs(SAEDs). Preferably the CBDV is used in combination with a SAED with amechanism of action which acts via sodium or calcium channels, morepreferably one which:

-   -   modifies low-threshold or transient neuronal calcium currents,        as exemplified by ethosuximide; or    -   reduces high-frequency neuronal firing and sodium-dependent        action potentials and may additionally enhance GABA effects, as        exemplified by valproate.

Alternatively the CBDV is used in combination with a SAED with amechanism of action which:

-   -   enhances GABAergic inhibition, as exemplified by phenobarbital.

BACKGROUND

Epilepsy is a chronic neurological disorder presenting a wide spectrumof diseases that affects approximately 50 million people worldwide(Sander, 2003). Advances in the understanding of the body's internal‘endocannabinoid’ system has lead to the suggestion that cannabis-basedmedicines may have the potential to treat this disorder ofhyperexcitability in the central nervous system (Mackie, 2006,Wingerchuk, 2004, Alger, 2006).

Cannabis has been ascribed both pro-convulsant (Brust et al., 1992) andanti-convulsant effects. Therefore, it remains to determine whethercannabinoids represent a yet to be unmasked therapeutic anticonvulsantor, conversely, a potential risk factor to recreational and medicinalusers of cannabis (Ferdinand et al., 2005).

In 1975 Consroe et al. described the case of young man whose standardtreatment (phenobarbital and phenytoin), didn't control his seizures.When he began to smoke cannabis socially he had no seizures. Howeverwhen he took only cannabis the seizures returned. They concluded that‘marihuana may possess an anti-convulsant effect in human epilepsy’.

A study by Ng (1990) involved a larger population of 308 epilepticpatients who had been admitted to hospital after their first seizure.They were compared to a control population of 294 patients who had nothad seizures, and it was found that using cannabis seemed to reduce thelikelihood of having a seizure. However this study was criticized in anInstitute of Medicine report (1999) which claimed it was ‘weak’, as thestudy did not include measures of health status prior to hospitaladmissions and differences in their health status might have influencedtheir drug use rather than the other way round.

Three controlled trials have investigated the anti-epilepsy potential ofcannabidiol. In each, cannabidiol was given in oral form to sufferers ofgeneralised grand mal or focal seizures.

Cunha et al (1980) reported a study on 16 grand mal patients who werenot doing well on conventional medication. They received their regularmedication and either 200-300 mg of cannabidiol or a placebo. Of thepatients who received CBD, 3 showed complete improvement, 2 partial, 2minor, while 1 remained unchanged. The only unwanted effect was mildsedation. Of the patients who received the placebo, 1 improved and 7remained unchanged.

Ames (1986) reported a less successful study in which 12 epilepticpatients were given 200-300 mg of cannabidiol per day, in addition tostandard antiepileptic drugs. There seemed to be no significantimprovement in seizure frequency.

Trembly et al (1990 performed an open trial with a single patient whowas given 900-1200 mg of cannabidiol a day for 10 months. Seizurefrequency was markedly reduced in this single patient.

In addition to the disclosures suggesting CBD may be beneficial there isa report (Davis & Ramsey) of tetrahydrocannabinol (THC) beingadministered to 5 institutionalized children who were not responding totheir standard treatment (phenobarbital and phenoytin). One becameentirely free of seizures, one became almost completely free ofseizures, and the other three did no worse than before.

In WO 2006/054057 it is suggested that the cannabinoidTetrahydrocannabivarin (THCV) may behave as anti epileptic, somethingconfirmed by Thomas et al 2005.

The application WO 2007/138322 shows CBD to be an inverse agonists atthe CB1 and CB2 receptors and suggests this compound and structurallyrelated compounds including CBDV, may have a therapeutic benefit in awide range of conditions which involve these receptors. Morespecifically the data demonstrates that the cannabinoid CBD reducedbodyweight in rats.

However other work on cannabinoids has shown that despite THCV'sstructural similarity to THC the two compounds behave quite differentlyat the CB1 receptor and consequently it does not follow that the propylcannabinoid analogs will behave as their pentyl equivalents.

In addition a study in 2007 by Deshpande et al. established that the CB1antagonist rimonabant was a pro-convulsant; this study demonstrated thatantagonism of the CB1 receptor caused epileptic activity. The inferencefrom this study is that cannabinoids which act as antagonists of the CB1receptor may not be useful as anti-convulsants; indeed they mayexacerbate such a condition.

WO 2009/007697 describes a THCV and CBD pharmaceutical formulation. Sucha formulation is suggested to be of use in many different types ofdiseases including epilepsy.

The application WO 2007/083098 describes the use of cannabis plantextracts with neuroprotective properties. Cannabinoid extractscontaining THC and CBD were shown to be more effective than their purecounterparts in this area of medicine.

The application WO 02/064109 describes a pharmaceutical formulationwhere the cannabinoids THC and CBD are used. The application goes on tostate that the propyl analogs of these cannabinoids may also be used inthe formulation. Since this application was written it has been shownthat THCV behaves in a very different manner to THC and therefore theassumption that the propyl analogs of cannabinoids may behave in asimilar manner to their pentyl counterparts is now not valid.

The application GB0911580.9 describes the use of THCV for the treatmentof generalised seizures, and also describes the use of CBD incombination with THCV.

However, there are more than forty recognisable types of epilepticsyndrome partly due to seizure susceptibility varying from patient topatient (McCormick and Contreras, 2001, Lutz, 2004) and a challenge isfinding drugs effective against these differing types.

Neuronal activity is a prerequisite for proper brain function. However,disturbing the excitatory—inhibitory equilibrium of neuronal activitymay induce epileptic seizures. These epileptic seizures can be groupedinto two basic categories:

a) partial, and

b) generalised seizures.

Partial seizures originate in specific brain regions and remainlocalised—most commonly the temporal lobes (containing the hippocampus),whereas generalised seizures appear in the entire forebrain as asecondary generalisation of a partial seizure (McCormick and Contreras,2001, Lutz, 2004). This concept of partial and generalised seizureclassification did not become common practice until the InternationalLeague Against Epilepsy published a classification scheme of epilepticseizures in 1969 (Merlis, 1970, Gastaut, 1970, Dreifuss et al., 1981).

The International League Against Epilepsy further classified partialseizures, separating them into simple and complex, depending on thepresence or the impairment of a consciousness state (Dreifuss et al.,1981).

The league also categorized generalised seizures into numerous clinicalseizure types, some examples of which are outlined below:

Absence seizures occur frequently, having a sudden onset andinterruption of ongoing activities. Additionally, speech is slowed orimpeded with seizures lasting only a few seconds (Dreifuss et al.,1981).

Tonic-clonic seizures, often known as “grand mal”, are the mostfrequently encountered of the generalised seizures (Dreifuss et al.,1981). This generalised seizure type has two stages: tonic musclecontractions which then give way to a clonic stage of convulsivemovements. The patient remains unconscious throughout the seizure andfor a variable period of time afterwards.

Atonic seizures, known as “drop attacks”, are the result of sudden lossof muscle tone to either a specific muscle, muscle group or all musclesin the body (Dreifuss et al., 1981).

The onset of epileptic seizures can be life threatening with sufferersalso experiencing long-term health implications (Lutz, 2004). Theseimplications may take many forms:

-   -   mental health problems (e.g. prevention of normal glutamatergic        synapse development in childhood);    -   cognitive deficits (e.g. diminishing ability of neuronal        circuits in the hippocampus to learn and store memories); and    -   morphological changes (e.g. selective loss of neurons in the CA1        and CA3 regions of the hippocampus in patients presenting mesial        temporal lobe epilepsy as a result of excitotoxicity) (Swann,        2004, Avoli et al., 2005)

It is noteworthy that epilepsy also greatly affects the lifestyle of thesufferer—potentially living in fear of consequential injury (e.g. headinjury) resulting from a grand mal seizure or the inability to performdaily tasks or the inability to drive a car unless having had a lengthyseizure-free period (Fisher et al., 2000).

Despite the historic work on CBD in epilepsy in the 1980's/1990'sresearch in the field of anti-convulsants has focused on many othercandidates many of which are now approved for use in the treatment ofepilepsy. Such drugs include: acetozolamide, carbamazepine, clobazam,clonazepam, ethosuximide, eslicarbazepine acetate, gabapentin,lacosamide, lamotriquine, levetiracetam, oxcarbazepine, Phenobarbital,phenytoin, pregabalin, primidone, rufinamide, sodium valproate,tiagabine, topiramate, valproate, vigabatrin, and zonisamide.

The mode of action of some of these is understood and for others isunknown. Some modes of action are set out in Table 1 below: (Adaptedfrom: Schachter S C. Treatment of seizures. In: Schachter S C, Schomer DL, eds. The comprehensive evaluation and treatment of epilepsy. SanDiego, Calif.: Academic Press; 1997. p. 61-74)

TABLE 1 Sodium or calcium or GABA channel Antiepileptic drug Mechanismof action involvement Barbiturates: Enhances GABAergic inhibition GABAprimidone (Mysoline), phenobarbital Carbamazepine Inhibitsvoltage-dependent sodium Sodium (Tegretol, channels Tegretol-XR,Carbatrol) Ethosuximide Modifies low-threshold or transient Calcium(Zarontin) neuronal calcium currents Felbamate (Felbatol) UnknownGabapentin Unknown (Neurontin) Lamotrigine Inhibits voltage-dependentsodium Sodium (Lamictal) channels, resulting in decreased release of theexcitatory neurotransmitters glutamate and aspartate Phenytoin Blockssodium-dependent action Sodium/ (Dilantin, Phenytek) potentials; reducesneuronal Calcium calcium uptake Valproate (Depakote, Reduceshigh-frequency neuronal Sodium/ Depakote ER, firing and sodium-dependentaction GABA Depakene, valproic potentials; enhances GABA effects acid)

However despite the introduction of some twenty different compounds fortreatment of epilepsy over the last twenty years there remains a needfor alternate drugs for several reasons:

-   -   i) 1-2% of the world's population suffer from epilepsy        (http://www.ncbi.nlm.nih.gov/sites/ppmc/articles/PMC1808496/);    -   ii) Of these 30% are refractory to existing treatments; and    -   iii) There are also notable motor side effects in the existing        therapies (http://en.wikipedia.org/wiki/Epilepsy).

For example valproate and ethosuximide both exhibit notable motor andother side effects (including sedation) when given to rats at dosesgreater than 200 mg/kg, as does phenobarbitone at doses greater than 250mg/kg in rat models of epilepsy.

Three well-established and extensively used in vivo models of epilepsyare:

-   -   pentylenetetrazole-induced (PTZ) model of generalised seizures        (Obay et al., 2007, Rauca et al., 2004);    -   pilocarpine-induced model of temporal lobe (i.e. hippocampus)        seizures (Pereira et al., 2007); and    -   penicillin-induced model of partial seizures (Bostanci and        Bagirici, 2006).

These provide a range of seizure and epilepsy models, essential fortherapeutic research in humans.

SUMMARY OF THE INVENTION

It is an object of the present invention to identify novelanti-convulsants for use in the treatment of epilepsy.

Preferably the novel anti-convulsant will be effective in areascurrently not adequately provided for by existing medications, standardanti-epileptic drugs (SAEDs).

Preferably the novel anti-convulsant will have a better side effectprofile than existing SAEDs particularly when it comes to motor sideeffects.

Additionally is would be desirable for the compounds to work alongsidestandard treatments for epilepsy, addressing unmet needs and/or allowinglower dosages to be used thereby countering some of the adverse effectsof such existing SAEDs.

According to one aspect of the invention, a formulation comprising CBDVand at least one pharmaceutically acceptable excipient.

According to another aspect of the invention, a method for the treatmentof epileptic seizures is provided. The method comprises administering toa subject in need thereof a therapeutically effective amount of thephytocannabinoid CBDV. In some embodiments, the type of epilepticseizure to be treated is a generalised seizure or a temporal lobeseizure. The CBDV may be administered with one or more therapeuticallyeffective phytocannabinoids. Examples of one or more therapeuticallyeffective phytocannabinoids include, but are not limited to THCV andCBD. In some embodiments, the CBDV is administered with one or moretherapeutically effective phytocannabinoids, wherein the one or moretherapeutically effective phytocannabinoids are THCV and CBD.

In some embodiments, the CBDV is in an isolated form. In someembodiments, the CBDV is in the form of a botanical drug substance.

The CBDV may be used in combination with a standard anti-epileptic drug.Examples of standard anti-epileptic drug includes, but are not limited,standard anti-epileptic drug having a mechanism of action which acts viasodium or calcium channels, or enhances GABAergic inhibition. In someembodiments, the standard anti-epileptic drug is phenobarbital. In someembodiments, the standard anti-epileptic drug having a mechanism ofaction which acts via sodium or calcium channels either modifieslow-threshold or transient neuronal calcium currents; or reduceshigh-frequency neuronal firing and sodium-dependent action potentialsand may additionally enhance GABA effects. In some embodiments, thestandard anti-epileptic drug is slected from the group comprisingethosuximide and valproate.

According to another aspect of the invention, a method for the treatmentof epileptic seizures is provided. The method comprises administering toa subject in need thereof a therapeutically effective amount of acannabis plant extract comprising a phytocannabinoid containingcomponent and a non-phytocannabinoid containing component, wherein thephytocannabinoid containing component comprises at least 50% (w/w) ofthe cannabis plant extract and contains as a principal phytocannabinoid,CBDV and as a secondary phytocannabinoid, CBD, and wherein thenon-phytocannabinoid containing component comprises a monoterpenefraction and a sesquiterpene fraction. In some embodiments, the methodfurther comprises administering of THCV.

In some embodiments, the phytocannabinoid containing component comprises64-78% (w/w) of the cannabis plant extract. In some embodiments, thephytocannabinoid containing component comprises 52-64% (w/w) CBDV of thetotal phytocannabinoid fraction. In some embodiments, thephytocannabinoid containing component comprises 22-27% (w/w) CBD of thetotal phytocannabinoid fraction. In In some embodiments, thephytocannabinoid containing component comprises 3.9-4.7% (w/w) THCV ofthe total phytocannabinoid fraction.

DEFINITIONS

Phytocannabinoids” are cannabinoids that originate from nature and canbe found in the cannabis plant. The phytocannabinoids can be isolatedcannabinoids or present as a botanical drug substance.

An “isolated cannabinoid” is defined as a phytocannabinoid that has beenextracted from the cannabis plant and purified to such an extent thatall the additional components such as secondary and minor cannabinoidsand the non-cannabinoid fraction have been removed.

A “botanical drug substance” or “BDS” is defined in the Guidance forIndustry Botanical Drug Products Draft Guidance, August 2000, USDepartment of Health and Human Services, Food and Drug AdministrationCentre for Drug Evaluation and Research as: “A drug derived from one ormore plants, algae, or microscopic fungi. It is prepared from botanicalraw materials by one or more of the following processes: pulverisation,decoction, expression, aqueous extraction, ethanolic extraction or othersimilar processes.” A botanical drug substance does not include a highlypurified or chemically modified substance derived from natural sources.Thus, in the case of cannabis, BDS derived from cannabis plants do notinclude highly purified Pharmacopoeial grade cannabinoids.

Preferably the components and the relative amounts thereof in the BDSare characterised to the extent that qualitatively at least 90% of thecomponents are identified.

In the present invention a BDS is considered to have two components: thephytocannabinoid-containing component and the non-phytocannabinoidcontaining component. Preferably the phytocannabinoid-containingcomponent is the larger component comprising greater than 50% (w/w) ofthe total BDS and the non-phytocannabinoid containing component is thesmaller component comprising less than 50% (w/w) of the total BDS.

The amount of phytocannabinoid-containing component in the BDS in someembodiments is greater than 55% (w/w), through 60%, 65%, 70%, 75%, 80%to 85% or more of the total extract. The actual amount is likely todepend on the starting material used and the method of extraction used.

The “principal phytocannabinoid” in a BDS is the phytocannabinoid thatis present in an amount that is higher than that of the otherphytocannabinoids. Preferably the principal phytocannabinoid is presentin an amount greater than 40% (w/w) of the total extract. Morepreferably the principal phytocannabinoid is present in an amountgreater than 50% (w/w) of the total extract. More preferably still theprincipal phytocannabinoid is present in an amount greater than 60%(w/w) of the total extract.

The amount of the principal phytocannabinoid in the BDS is preferablygreater than 75% of the phytocannabinoid-containing fraction, morepreferably still greater than 85% of the phytocannabinoid-containingfraction, and more preferably still greater than 95% of thephytocannabinoid-containing fraction.

In some cases, such as where the principal cannabinoid is either CBDV orTHCVA the amount of the principal phytocannabinoid in the BDS is lower.Here the amount of phytocannabinoid is preferably greater than 55% ofthe phytocannabinoid-containing fraction.

The “secondary phytocannabinoid/s” in a BDS is the phytocannabinoid/sthat is/are present in significant proportions. Preferably the secondaryphytocannabinoid is present in an amount greater than 5% (w/w) of thetotal extract, more preferably greater than 10% (w/w) of the totalextract, more preferably still greater than 15% (w/w) of the totalextract. Some BDS's will have two or more secondary phytocannabinoidsthat are present in significant amounts. However not all BDS's will havea secondary phytocannabinoid. For example CBG BDS does not have asecondary phytocannabinoid in its extract.

The “minor phytocannabinoid/s” in a BDS can be described as theremainder of all the phytocannabinoid components once the principal andsecondary phytocannabinoids are accounted for. Preferably the minorphytocannabinoids are present in total in an amount of less than 10%(w/w) of the total extract, more preferably still less than 5% (w/w) ofthe total extract, and most preferably the minor phytocannabinoid ispresent in an amount less than 2% (w/w) of the total extract.

Typically the non-phytocannabinoid containing component of the BDScomprises terpenes, sterols, triglycerides, alkanes, squalenes,tocopherols and carotenoids.

These compounds may play an important role in the pharmacology of theBDS either alone or in combination with the phytocannabinoid.

The “terpene fraction” may be of significance and can be broken down bythe type of terpene: monoterpene or sesquiterpene. These terpenecomponents can be further defined in a similar manner to thecannabinoids.

The amount of non-phytocannabinoid containing component in the BDS maybe less than 45%, through 40%, 35%, 30%, 25%, 20% to 15% or less of thetotal extract. The actual amount is likely to depend on the startingmaterial used and the method of extraction used.

The “principal monoterpene/s” in a BDS is the monoterpene that ispresent in an amount that is higher than that of the other monoterpenes.Preferably the principal monoterpene/s is present in an amount greaterthan 20% (w/w) of the total terpene content. More preferably theprincipal monoterpene is present in an amount greater than 30% (w/w) ofthe total terpene content, more preferably still greater than 40% (w/w)of the total terpene content, and more preferably still greater than 50%(w/w) of the total terpene content. The principal monoterpene ispreferably a myrcene or pinene. In some cases there may be two principalmonoterpenes. Where this is the case the principal monoterpenes arepreferably a pinene and/or a myrcene.

The “principal sesquiterpene” in a BDS is the sesquiterpene that ispresent in an amount that is higher than all the other terpenes.Preferably the principal sesquiterpene is present in an amount greaterthan 20% (w/w) of the total terpene content, more preferably stillgreater than 30% (w/w) of the total terpene content. The principalsesquiterpene is preferably a caryophyllene and/or a humulene.

The sesquiterpene components may have a “secondary sesquiterpene”. Thesecondary monoterpene is preferably a pinene, which is preferablypresent at an amount greater than 5% (w/w) of the total terpene content,more preferably the secondary terpene is present at an amount greaterthan 10% (w/w) of the total terpene content.

The secondary sesquiterpene is preferably a humulene which is preferablypresent at an amount greater than 5% (w/w) of the total terpene content,more preferably the secondary terpene is present at an amount greaterthan 10% (w/w) of the total terpene content.

Alternatively botanical extracts may be prepared by introducing isolatedphytocannabinoids into a non-cannabinoid plant fraction as can beobtained from a zero cannabinoid plant or a CBG-free BDS.

The structure of CBDV is as shown below:

CBDV Cannabidivarin

Phytocannabinoids can be found as either the neutral (decarboxylatedform) or the carboxylic acid form depending on the method used toextract the cannabinoids. For example it is known that heating thecarboxylic acid form will cause most of the carboxylic acid form todecarboxylate into the neutral form.

Phytocannabinoids can also occur as either the pentyl (5 carbon atoms)or propyl (3 carbon atoms) variant. Initially it was thought that thepropyl and pentyl variants would have similar properties, however recentresearch suggests this is not true. For example the phytocannabinoid THCis known to be a CB1 receptor agonist whereas the propyl variant THCVhas been discovered to be a CB1 receptor antagonist meaning that it hasalmost opposite effects.

This is confirmed by Pertwee (2000) in Cannabinoid receptor ligands:clinical and neuropharmacological considerations relevant to future drugdiscovery and development, which describes potential therapeutic targetsfor CB1 receptor antagonists which include appetite suppression, thereduction of L-dopa dyskinesia in patient's with Parkinson's disease,management of acute schizophrenia and the amelioration of cognitivememory dysfunctions associated with Alzheimer's disease. All of thesetherapeutic targets are very different from those suggested for CB1receptor agonists such as appetite stimulation and reduction of pain.

A preferred CBDV formulation is one comprising CBDV BDS, isolated CBDVor synthetic CBDV in a form suitable for oral delivery. Forms suitablefor oral delivery include capsules and tablets. In such formulations theactive may be mixed with one or more excipients and may additionallycomprise one or more SAEDs, although more typically combinations withSAEDs would be administered separately, sequentially or simultaneously.

Unit dosage amounts may vary depending on the type and severity of theepilepsy to be treated. Each dosage unit may comprise less than or equalto 1000 mg of CBDV and the number of doses to be taken may also bevaried to suit a patient's requirements.

BRIEF SUMMARY OF THE DISCLOSURE

In accordance with a first aspect of the present invention there isprovided a phytocannabinoid CBDV for use in the treatment of epilepticseizures.

Surprisingly in an MES model of epilepsy CBDV showed much greateranti-convulsant activity than CBD.

In accordance with a second aspect of the present invention there isprovided the use of the phytocannabinoid CBDV in the manufacture of amedicament for use in the treatment of epileptic seizures.

The medicament may be a formulation comprising CBDV and at least onepharmaceutically acceptable excipient.

In accordance with a third aspect of the present invention there isprovided a method for the treatment of epileptic seizures, whichcomprises administering to a subject in need thereof a therapeuticallyeffective amount of the phytocannabinoid CBDV.

Preferably the type of epileptic seizure to be treated is a generalisedseizure or a temporal lobe seizure.

In one embodiment the CBDV is used in combination with one or moretherapeutically effective phytocannabinoids.

Preferably the one or more therapeutically effective phytocannabinoid isTHCV and/or CBD.

In one embodiment the CBDV is in an isolated form.

In a further embodiment the CBDV is in the form of a botanical drugsubstance.

In a further embodiment still, the CBDV is used in combination with astandard anti-epileptic drug. The SAED may be one with a mechanism ofaction which acts via sodium or calcium channels, more preferably onewhich:

-   -   modifies low-threshold or transient neuronal calcium currents,        as exemplified by ethosuximide; or    -   reduces high-frequency neuronal firing and sodium-dependent        action potentials and may additionally enhance GABA effects, as        exemplified by valproate.

Alternatively the SAED may be one with a mechanism of action whichenhances GABAergic inhibition, as exemplified by phenobarbital.

The combination (see Examples) demonstrates combinations can:

-   -   a. reduce the incidence of tonic-clonic seizures;    -   b. increase the amount of time a patient is seizure free;    -   c. increase the latency to onset of seizure;    -   d. decrease the overall duration of the seizure;    -   e. reduce the severity and mortality of the seizures; and    -   f. reduce the motor and other side effects (including sedation)        associated with the SAEDs.

Thus, the combinations are particularly well suited in the treatment ofconditions generally considered refractory to existing medication. Thecombinations would also appear to allow for the use of lower doses ofthe SAED's than would be used were the SAED to be used alone.

In accordance with a fourth aspect of the present invention there isprovided a cannabis plant extract comprising a phytocannabinoidcontaining component and a non-phytocannabinoid containing component,wherein the phytocannabinoid containing component comprises at least 50%(w/w) of the cannabis plant extract and contains as a principalphytocannabinoid, CBDV, and as a secondary phytocannabinoid, CBD, andwherein the non-phytocannabinoid containing component comprises amonoterpene fraction and a sesquiterpene fraction, for use in thetreatment of epileptic seizures.

In accordance with a fifth aspect of the present invention there isprovided the use of a cannabis plant extract comprising aphytocannabinoid containing component and a non-phytocannabinoidcontaining component, wherein the phytocannabinoid containing componentcomprises at least 50% (w/w) of the cannabis plant extract and containsas a principal phytocannabinoid, CBDV, and as a secondaryphytocannabinoid, CBD, and wherein the non-phytocannabinoid containingcomponent comprises a monoterpene fraction and a sesquiterpene fraction,in the manufacture of a medicament for use in the treatment of epilepticseizures.

Preferably the cannabis plant extract further comprises THCV.

Preferably the phytocannabinoid containing component comprises 64-78%(w/w) of the cannabis plant extract.

Preferably the phytocannabinoid containing component comprises 52-64%(w/w) CBDV of the total phytocannabinoid fraction, 22-27% (w/w) CBD ofthe total phytocannabinoid fraction and 3.9-4.7% (w/w) THCV of the totalphytocannabinoid fraction.

BRIEF DESCRIPTION OF THE DRAWINGS

Embodiments of the invention are further described hereinafter withreference to the accompanying drawings, in which

FIG. 1 shows the effect of CBDV on onset and development of PTZ-inducedseizures;

FIG. 2 shows the effects of CBDV on seizure severity and mortality;

FIG. 3 A-C shows the effect of CBDV and ethosuximide on PTZ-inducedseizures;

FIG. 4 A-D shows the effect of CBDV and ethosuximide on incidence ofseizures and mortality in PTZ-induced seizures;

FIG. 5 shows the effect of CBDV and valproate on PTZ-induced seizures(onset latency and seizure duration);

FIG. 6 A-B shows the effect of CBDV and valproate on seizure severityand mortality in PTZ-induced seizures;

FIG. 7 A-D shows the effect of different doses of CBDV alone inPilocarpine-induced seizures (seizure severity, mortality, seizure freeand onset latency);

FIG. 8 A-D shows the effect of different doses of CBDV on seizureepisodes in Pilocarpine-induced seizures (number of episodes, episodeseverity, episode latency and episode duration);

FIG. 9 A-B shows the effect of high dose (200 mg/Kg) CBDV and valproatein Pilocarpine-induced seizures (severity and mortality);

FIG. 10 A-B shows the effect of high dose (200 mg/Kg) CBDV and valproatein Pilocarpine-induced seizures (bilateral latency and incidence);

FIG. 11A-B shows the effect of high dose (200 mg/Kg) CBDV and valproatein Pilocarpine-induced seizures (tonic clonic incidence and duration);

FIG. 12 A-B shows the effect of CBDV and phenobarital inPilocarpine-induced seizures (severity and mortality);

FIG. 13 A-B shows the effect of CBDV and phenobarital inPilocarpine-induced seizures (seizure free and onset latency);

FIG. 14 shows the effects of THCV BDS and 70 mg/kg PTZ on latencies toinitial and later seizure severities;

FIG. 15 shows the effects of THCV BDS and 70 mg/kg PTZ on seizureduration and time to death;

FIG. 16 shows the effects of THCV BDS and 70 mg/kg PTZ on medianseverity scores;

FIG. 17 shows the effects of THCV BDS and 70 mg/kg PTZ on mortalityrates;

FIG. 18 shows the effects of THCV BDS and 80 mg/kg PTZ on latencies toinitial and later seizure severities;

FIG. 19 shows the effects of THCV BDS and 80 mg/kg PTZ on seizureduration and time to death;

FIG. 20 shows the effects of THCV BDS and 80 mg/kg PTZ on medianseverity scores;

FIG. 21 shows the effects of THCV BDS and 80 mg/kg PTZ on mortalityrates;

FIG. 22 A-D show PTZ-induced seizure development and duration withisolated THCV;

FIG. 23 A-B show the effect of CBD on PTZ-induced seizures;

FIG. 24 shows the effect of vehicle on rotarod performance; and

FIG. 25 shows the effect of CBDV on rotarod performance.

Legend to FIG. 1: A-C: mean latency to seizure onset (A), clonic (B) andtonic-clonic (C) seizures in s. Statistical significance was assessed byANOVA and post hoc Tukey test, 1.30.05 was considered to be significantin both cases. Data is presented ±S.E.M., * indicates p<0.05.

Legend to FIG. 2: A: Median severity of seizures (grey line), also shownis the 25^(th) and 75^(th) percentiles (black horizontal lines) and themaximum and minimum values (upward and downward error barsrespectively). B: Proportion of animals in each group that developedtonic-clonic seizures. C: Proportion of animals in each group that died.D: Proportion of animals in each group that remained seizure free afterPTZ administration. *, ** and *** indicate p<0.05, 0.01 and 0.001respectively. A: median data tested by ANOVA and post hoc Tukey's test.B-D: Percentages tested by binomial statistics test.

Legend to FIG. 3: A: onset latency ±S.E.M. B: Severity; median valuesare shown in red, 25th and 75th percentiles are represented by boxes andmaxima and minima in each group by error bars. C: Seizure duration±S.E.M.

Legend to FIG. 4: A: Effects of CBDV on the proportion of animals thatremained seizure-free (%). B&C: Effects of CBDV on the proportion ofanimals that developed clonic (B) and tonic-clonic (C) seizures (%). D:Effects of CBDV on mortality (%).

Legend to FIG. 7: A: Effect of CBDV on overall seizure severity. Greylines indicate median severity for each group, “boxes” represent 25thand 75th percentile ranges, error bars represent maxima and minima. B,C: Effect of CBDV on percentage mortality (B) and the percentage ofanimals that remained seizure-free (C). Seizure-free was considered tobe a score of [1] or [0]. D: Onset latency (±S.E.M.) in seconds to firstdisplay of seizure severity [2] or above.

Legend to FIG. 8: A: the mean number of seizure episodes (per animal,only animals that experienced seizures were included). B: Medianseverity of all episodes in an experimental group, see FIG. 1 (PILO) fordescription of plot. C: Latency to 1st episode (±S.E.M.) in seconds. D:Mean duration of all episodes in an experimental group (±S.E.M.).

Legend to FIG. 14: The mean latencies to first myoclonic jerk (FMJ) andscores of 3.5 are shown ±S.E.M. ∫=8-10.

Legend to FIG. 15: The mean durations of seizures in animals thatsurvived, and the time from first seizure sign to death in those thatdied, are shown ±S.E.M. for vehicle or for low, medium or high dosesn=3-10 dependent on proportions of animals that died within experimentalgroups. ∫=vehicle group had no deaths and so no value is shown here.

Legend to FIG. 16: Median severity scores for groups of animals treatedwith vehicle or with low, medium or high doses n=10 for all groups.

Legend FIG. 17: Mortality rates expressed as percentages for animalstreated with vehicle or with low, medium or high doses. n=10 for allgroups. ∫=vehicle group had no deaths, therefore no value is shown.

Legend to FIG. 18: The mean latencies to first myoclonic jerk (FMJ) andscores of 3.5 are shown ±S.E.M. for vehicle or for low, medium or highdoses. n=7-10.

Legend to FIG. 19: The mean durations of seizures in animals thatsurvived, and the time from first seizure sign to death in those thatdied, are shown ±S.E.M. for vehicle or for low, medium or high doses.n=3-7 dependent on proportions of animals that died within experimentalgroups.

Legend to FIG. 20: Median severity scores for groups of animals treatedwith vehicle or with low, medium or high doses. n=10 for all groups.

Legend to FIG. 21: Mortality rates expressed as percentages for animalstreated with vehicle or with low, medium or high doses. n=10 for allgroups.

Legend to FIG. 22: A, B and C show the mean latency (s) from injectionof 80 mg/kg PTZ to: first sign of seizure (A); development of myoclonicseizures (B) and full tonic-clonic seizures (C) for vehicle andTHCV-dosed groups. n=5-16 depending on incidence of each marker within aspecific group). D shows the mean duration of seizures (s) in animalsthat survived post-seizure. All values ±S.E.M., * indicates significantdifference from vehicle group (P<0.05; Mann-Whitney U test).

Legend to FIG. 23: A: % mortality experienced as a result of IPinjection of 80 mg/kg PTZ in vehicle and CBD-dosed (1, 10,100 mg/kg CBD)animals (n=15 for all groups). B: % of vehicle- and CBD-dosed (1, 10,100mg/kg CBD) animals that experienced tonic-clonic seizures as a result ofIP injection of 80 mg/kg PTZ. * indicates significant result (p<0.01).

Legend to FIG. 24: Median latency to falling ±S.E. from the rotarodfollowing administration of saline and 2:1:17 cremaphor:ethanol:saline.

Legend to FIG. 25: Median latency to falling from rotarod (grey bars)with 25^(th) and 75^(th) percentiles (black boxes) and maximum andminimum values (error bars) also presented.

DETAILED DESCRIPTION

Examples 1 to 5 below describe the use of isolated CBDV in differentmodels of epilepsy. Further examples will describe phytocannabinoid BDSwhich comprise along with the principal cannabinoid other secondary andminor cannabinoids along with a non-phytocannabinoid containingfraction.

Example 1 Use of Isolated CBDV in Two In Vitro Epileptiform Models inHippocampal Brain Slices

Hippocampal slices were produced acutely from P>21 Wistar rats andactivity recorded by multi-electrode arrays (MEA).

To induce epileptiform activity, either Mg²⁺ was removed (Mg²⁺-freemodel) or 100 μM 4-aminopyridine was added (4-AP model). 30 min afterepileptiform burst activity was established, CBDV was added cumulatively(1, 10, 100 μM; 30 min each).

The effects of CBDV on epileptiform burst amplitude and duration weremeasured (Table 2.1).

Overall, CBDV at ≧10 μM or 100 μM significantly decreased burst durationand amplitude in both models with CA1 and DG regions most sensitive andCA3 least sensitive to the anti-epileptiform effects of CBDV.

TABLE 2.1 Effects of CBDV on epileptiform activity induced in the Mg²⁺free and 4-AP models CBDV Burst amplitude (% of control) Burst duration(% of control) (μM) DG CA3 CA1 DG CA3 CA1 Mg²⁺ _(o)- 1 89.8 ± 8.6 112.7± 13.7 82.23 ± 10.4 90.6 ± 4.5  101.7 ± 2.3  99.3 ± 4.5  free 10  86.4 ±3.6* 104.8 ± 10.3  79.9 ± 6.9** 92.0 ± 3.2*  93.9 ± 4.1  91.2 ± 3.8*model 100  79.5 ± 5.6** 102.9 ± 13.0  80.4 ± 8.0* 75.6 ± 5.4** 78.5 ±6.6*  74.0 ± 5.8** 4-AP 1 94.2 ± 3.0 103.0 ± 5.8  89.3 ± 5.6 95.7 ± 5.9 91.0 ± 6.0  104.3 ± 8.0  model 10 91.2 ± 4.9 121.9 ± 17.0 88.3 ± 5.283.8 ± 4.4** 82.5 ± 4.8* 85.9 ± 6.2* 100 95.9 ± 4.3 110.3 ± 7.0   89.5 ±5.3* 83.4 ± 4.1** 79.7 ± 5.4* 85.9 ± 5.8* Data is mean ± S.E.M; *= p ≦0.05 and **= p ≦ 0.01 respectively, Wilcoxon paired test. Data from 5rats/model, n = 9-13 electrodes.

Example 2 Use of Isolated CBDV in the PTZ Model of Generalised SeizuresMethodology: Animals:

Male Wistar rats (P24-29; 75-110 g) were used to assess the effects ofthe phytocannabinoid CBDV in the PTZ model of generalised seizures.Animals were habituated to the test environment, cages, injectionprotocol and handling prior to experimentation. Animals were housed in aroom at 21° C. on a 12 hour light: dark cycle (lights on 0900) in 50%humidity, with free access to food and water.

The human dose equivalent (HED) can be estimated using the followingformula:

${HED} = {{Animal}\mspace{14mu} {dose}\mspace{14mu} \left( {{mg}\text{/}{kg}} \right)\mspace{14mu} {multiplied}\mspace{14mu} {by}\frac{{Animal}\mspace{14mu} K_{m}}{{Human}\mspace{14mu} K_{m}}}$

The K_(m) for a rat is 6 and the K_(m) for a human is 37.Thus, for a human of approx 60 Kg a 200 mg/Kg dose in rat would equateto a human daily dose of about 2000 mg.

Experimental Setup:

Five 6 L Perspex tanks with lids were placed on a single bench withdividers between them. Closed-circuit television (CCTV) cameras weremounted onto the dividers to observe rat behaviour. Sony Topica CCDcameras (Bluecherry, USA) were linked via BNC cables to a low-noise PCvia Brooktree digital capture cards (Bluecherry, USA). Zoneminder(http://www.zoneminder.com) software was used to monitor rats, start andend recordings and manage video files. In-house Linux scripts were usedto encode video files into a suitable format for further offlineanalysis using The Observer (Noldus Technologies).

PTZ Model:

A range of doses of PTZ (50-100 mg/kg body weight) were used todetermine the best dose for induction of seizures (see below). As aresult, a dose of 80 mg/kg injected intra-peritoneally (IP; stocksolution 50 mg/ml in 0.9% saline) were used to screen the CBDV.

Experimental Protocols:

On the day of testing, pure CBDV was administered via intra-peritoneal(i.p.) injection at doses of 50, 100 and 200 mg/kg alongside animalsthat were injected with a matched volume of the cannabinoid vehicle(2:1:17 ethanol:Cremophor: 0.9% w/v NaCI solution), which served as thenegative control group. Animals were then observed for 1 hour, afterwhich time they received an IP injection of 80 mg/kg PTZ. Negativevehicle controls were performed in parallel with cannabinoid-dosedsubjects. After receiving a dose of PTZ, animals were observed andvideoed to determine the severity of seizure and latency to severalseizure behaviour types (see in vivo analysis, below). Animals werefilmed for half an hour after last sign of seizure, and then returned totheir cage.

In vivo Analysis:

Animals were observed during experimental procedures, but all analysiswas performed offline on recorded video files using The Observerbehavioural analysis software (Noldus, Netherlands). A seizure severityscoring system was used to determine the levels of seizure experiencedby subjects (Pohl & Mares, 1987). All signs of seizure were detailed forall animals.

TABLE 3.1 Seizure severity scoring scale, adapted from Pohl & Mares,1987. Seizure Righting score Behavioural expression reflex 0 No changesto behaviour Preserved 0.5 Abnormal behaviour (sniffing, excessivewashing, Preserved orientation) 1 Isolated myoclonic jerks Preserved 2Atypical clonic seizure Preserved 3 Fully developed bilateral forelimbclonus Preserved 3.5 Forelimb clonus with tonic component and bodyPreserved twist 4 Tonic-clonic seizure with suppressed tonic phase Lost5 Fully developed tonic-clonic seizure Lost 6 DeathLatency from Injection of PTZ to Specific Indicators of SeizureDevelopment:

The latency (in seconds) from injection of PTZ to first myoclonic jerk(FMJ; score of 1), and to the animal attaining “forelimb clonus withtonic component and body twist” (score of 3.5) were recorded. FMJ is anindicator of the onset of seizure activity, whilst >90% of animalsdeveloped scores of 3.5, and so is a good marker of the development ofmore severe seizures. Data are presented as the mean±S.E.M. within anexperimental group.

Maximum Seizure Severity:

This is given as the median value for each experimental group based onthe scoring scale below.

% Mortality:

The percentage of animals within an experimental group that died as aresult of PTZ-induced seizures. Note that the majority of animals thatdeveloped tonic-clonic seizures (scores of 4 and 5) died as a result,and that a score of 6 (death) automatically denotes that the animal alsoexperienced tonic-clonic seizures.

Seizure Duration:

The time (in seconds) from the first sign of seizure (typically FMJ) toeither the last sign of seizure or, in the case of subjects that died,the time of death—separated into animals that survived and those thatdid not. This is given as the mean±S.E.M. for each experimental group.

Statistics:

For measures of latency and severity, one way analysis of variance(ANOVA) was performed on the four groups together (vehicle and 50, 100and 200 mg/kg CBDV) to detect overall effects of CBDV (p≦30.05considered significant).

Significant ANOVA results were followed by post hoc tests to testdifferences between vehicle and drug groups (Tukey's test, p≦0.05considered significant).

Results:

FIG. 1 illustrates the onset and development of seizures by showing thelatency from administration of 80 mg/kg PTZ to: the onset of seizure(FIG. 1A); the development of clonic seizures (FIG. 1B) and thedevelopment of tonic-clonic seizures (FIG. 1C).

A significant effect of CBDV on the latency to seizure onset wasobserved (p=0.041; FIG. 1A); this measure was significantly higher inanimals that received 200 mg/kg CBDV than those that received vehiclealone (p=0.03).

A near-significant (p=0.055) effect of CBDV on latency to clonicseizures was observed (FIG. 1B), highlighting a significant increase inanimals administered 200 mg/kg CBDV compared to vehicle-treated animals(p=0.032).

No significant effect of CBDV on latency to tonic-clonic seizuresoverall or at any specific dose was observed (FIG. 1C) in spite of alarge difference in mean value between vehicle and 200 mg/kg CBDVgroups; this is likely to be due to the low number of animals treatedwith 200 mg/kg CBDV that developed these seizures

The severity of seizures experienced by animals in the different groupswas also assessed using four measures: median severity (FIG. 2A);proportion of animals that had tonic-clonic seizures (the most severeseizure type; FIG. 2B); the percentage mortality (FIG. 2C) and finallythe proportion of animals that remained seizure free after PTZadministration (FIG. 2D).

There was an overall significant effect of CBDV on seizure severity(p=0.007; FIG. 2A); animals treated with 200 mg/kg CBDV had asignificantly lower median severity than those treated with vehiclealone (p=0.014).

This was reflected in a lower proportion of animals treated with 200mg/kg CBDV reaching the most severe (tonic-clonic) seizures (3 of 15)compared to vehicle-treated animals (8 of 15; FIG. 2B; p=0.01).

This significant effect was maintained in animals treated with 100 mg/kgCBDV (4 of 15 tonic-clonic seizures; p=0.036), but not 50 mg/kg.

A significantly lower proportion of animals treated with 100 and 200mg/kg CBDV (1 and 2 out of 15 respectively) died compared to thevehicle-treated group (8 of 15; p=0.002 and <0.001 respectively; FIG.2C).

Finally, a significantly higher percentage of animals treated with 200mg/kg CBDV experienced no seizure at all (5 of 15) compared to thevehicle group (1 of 15; p=0.003; FIG. 2D).

Conclusion:

From the above data it would appear that CBDV shows great potential asan anti-epileptic drug.

Example 3 Use of Isolated CBDV with Standard Anti-Epileptic Drugs(SAEDs) in the PTZ Model of Generalised Seizures Methodology:

As described in Example 2 above. Varying doses of the SAED'sethosuximide and valproate were tested in combination to isolated CBDVat a dose of 200 mg/kg.

Results:

FIGS. 3 and 4 detail the use of the SAED ethosuximide (a drug operatingvia calcium channels) with isolated CBDV. Although the combination ofthe two compounds increased the onset latency, reduced seizure duration,resulted in more seizure free animals and reduced tonic/clonic seizuresthere was no statistically significant interaction between the twocompounds. The CBDV gave similar results to valproate for mortality,however significantly CBDV was able to lessen the severity of theepilepsy to a greater degree than the existing epileptic drug valproate.

FIGS. 5 and 6 detail the use of the SAED valproate (a drug operating viasodium channels) with isolated CBDV. When co-administered, valproate andCBDV independently induced significant decreases in onset latency,seizure severity and mortality although no synergistic effects ofcombinatorial administration were noted.

Both sets of results indicate benefits in their use in combination.

Example 4 Use of Isolated CBDV in the Pilocarpine Model of EpilepsyMethodology:

Isolated CBDV was injected intra-peritoneally (IP) in the standardvehicle (1:1:18 ethanol:Cremophor:0.9% ^(w)/_(v) NaCI) at doses of 50,100 and 200 mg/kg alongside animals that received vehicle alone at amatched volume. 15 minutes later methylscopolamine (1 mg/kg; to reduceperipheral muscarinic effects of pilocarpine) was administered followed,45 minutes later by pilocarpine (380 mg/kg, IP) administration.

Results:

FIGS. 7 and 8 details the effect of CBDV on pilocarpine-inducedseizures. As can be observed the lower doses of CBDV (50 and 100 mg/kg)decreased the mortality.

Example 5 Use of Isolated CBDV with Standard Anti-Epileptic Drugs(SAEDs) in the Pilocarpine Model of Epilepsy Methodology:

As described in Example 4 above, the SAEDs valproate and phenobarbitalwere used at various doses along with isolated CBDV at 200 mg/kg. Thesetwo drugs are representative of two classes of anti-convulsants whichhave different mechanisms of action. Valproate operates via sodiumchannels and Phenobarbital enhances GABAergic inhibition.

Results:

FIG. 9 details the data obtained when CBDV was used in combination withthe SAED valproate. Both CBDV and valproate exerted independent andpositive effects upon seizure severity although only CBDV and notvalproate independently caused a significant decrease in mortality. Thecombination of CBDV with valproate increased the seizure latency anddecreased the seizure incidence. However these data were notstatistically significant.

FIG. 10 details further data obtained when CBDV was used in combinationwith the SAED valproate. It shows that bilateral seizure incidence wassignificantly decreased by CBDV (particularly with the high doseValproate (250 mg/kg).

FIG. 11 details further data obtained when CBDV was used in combinationwith the SAED valproate. It shows that both tonic/clonic incidence andtotal tonic clonic duration decreased when CBDV was used in combinationwith all doses of Valporate and that the CBDV interaction (alone) withclonic tonic seizure was statistically significant.

FIG. 12 details the data obtained when CBDV was used in combination withthe SAED phenobarbital. As can be seen the CBDV significantly decreasesseverity and the combination is also significant.

FIG. 13 details further data obtained when CBDV was used in combinationwith the SAED phenobarbital. Although the data did not demonstratestatistical significance there was a strong trend, particularly at thelower dose levels of Phenobarbital, towards an increase in seizure freeanimals and increased onset latency.

Example 6 Analysis of Cannabinoid Botanical Drug Substances

As described in the following example, CBDV BDS comprises, as well asCBDV, the cannabinoids CBD and THCV. Given the finding disclosed inGB0911580.9 that CBD and THCV exhibit anti-convulsant activity, a CBDVextract containing in addition to CBDV, CBD and THCV make it potentiallymore interesting than isolated CBDV, particularly as extracts may onlypossess very low amounts of THC.

Cannabidivarin (CBDV) Botanical Drug Substance Analysis

A CBDV BDS can be obtained from extraction of CBDV-rich plants. Suchchemovars are bred specifically to produce a significant proportion oftheir cannabinoids as CBDV.

CBDV BDS can also be prepared by adding isolated CBDV to a cannabinoidfree BDS. Such a cannabinoid free BDS can be prepared from either a CBGBDS or a zero cannabinoid plant such as USO-31. Because CBG is the majorcannabinoid present in CBG BDS it is possible to remove the CBG presentrelatively easily using standard techniques known in the art such ascolumn chromatography. It is possible to fractionate the BDS completelyso that individual compounds can be removed for purification and theremainder recombined to produce, following solvent removal, a BDS freeof the selected compound(s).

The CBDV chemotype results from the breeding of plants which carry bothpostulated B_(D) and A_(PR) genes.

The B_(D) gene instruct the plants to synthesize the cyclic part of theCBD molecule and the A_(PR) gene instructs the plant to synthesize thismolecule with a propyl side chain, as opposed to the usual pentyl chainfound in CBD.

A CBDV chemovar has been bred and the BDS analysed as described in Table4.1 below:

TABLE 4.1 Cannabidivarin BDS amount in total and range Amount RangeRange Range CBDV BDS (% w/w) (±10%) (±25%) (±50%) CBDVA 0.14 0.13-0.150.11-0.18 0.07-0.21 CBDV 41.19 37.07-45.31 30.89-51.49 20.60-61.79 CBDA0.07 0.06-0.08 0.05-0.09 0.04-0.11 CBG 0.59 0.53-0.65 0.44-0.740.30-0.89 CBD 17.70 15.93-19.47 13.28-22.13 8.85-26.55 THCV 3.062.75-6.12 2.30-3.83 1.53-4.59 CBCV 4.35 3.92-4.79 3.26-5.44 2.18-6.53THC 0.88 0.79-0.97 0.66-1.10 0.44-1.32 CBDV (related 2.20 1.98-2.421.65-2.75 1.10-3.30 substances) CBC 0.93 0.84-1.02 0.70-1.16 0.47-1.40Total 71.11 Cannabinoids Total Non- 28.89 cannabinoids

The total phytocannabinoid containing fraction of CBDV BDS comprisesapproximately 41% of the total BDS. According to variation this fractionmay vary by ±10% up to ±50%.

TABLE 4.2 Cannabidivarin BDS by percentage cannabinoid Amount CBDV BDS(% of total cannabinoid) CBDVA 0.20 CBDV 57.92 CBDA 0.10 CBG 0.83 CBD24.89 THCV 4.30 CBCV 6.12 THC 1.24 CBDV (related 3.09 substances) CBC1.31

The amount of the principal phytocannabinoid in the CBDV BDS as apercentage of the phytocannabinoid containing fraction is approximately58%. According to variation this fraction may vary by ±10% up to ±50%.

In this Example it is intended that references be made to the principalor secondary components independently of the ‘other’ cannabinoids.

The finding that the CBDV BDS comprises the known anti-epilepticphytocannabinoids CBD and THCV in relatively large amounts andrelatively little THC, as compared to THCV extract below infers that theuse of CBDV in the form of a BDS will be a promising new treatment forepilepsy.

Tetrahydrocannabivarin (THCV) Botanical Drug Substance Analysis

Table 4.3 below details the cannabinoid components of THCV BDS, as canbe seen the secondary cannabinoid is THC and is present at a significantamount in comparison to the other cannabinoids.

TABLE 4.3 Tetrahydrocannabivarin BDS amount in total and range AmountRange Range Range THCV BDS (% w/w) (±10%) (±25%) (±50%) CBGV 0.150.14-0.17 0.11-0.19 0.07-0.23 CBNV 1.30 1.20-1.40 1.00-1.60 0.65-1.95THCV 64.49 58.04-70.94 48.37-80.61 32.25-96.74 CBCV 0.65 0.59-0.720.49-0.81 0.33-0.98 THC-C4 0.82 0.74-0.90 0.62-1.03 0.41-1.23 CBN 0.150.14-0.17 0.11-0.19 0.07-0.23 THCVA 0.36 0.32-0.40 0.27-0.45 0.18-0.54THC 13.43 12.09-14.77 10.07-16.79  7.72-20.15 Unknowns 0.58 0.52-0.640.44-0.73 0.29-0.87 Total 81.93 Cannabinoids Total Non- 18.07cannabinoidsThe total phytocannabinoid containing fraction of THCV BDS comprisesapproximately 74-90% (w/w) of the total BDS.

TABLE 4.4 Tetrahydrocannabivarin BDS by percentage cannabinoid AmountTHCV BDS (% of total cannabinoid) CBGV 0.18 CBNV 1.59 THCV 78.71 CBCV0.79 THC-C4 1.00 CBN 0.18 THCVA 0.44 THC 16.39 Unknowns 0.71

The amount of the principal phytocannabinoid in the THCV BDS as apercentage of the phytocannabinoid containing fraction is approximately71-87% (w/w). The THCV BDS also has a secondary cannabinoid THC which ispresent at approximately 14.8-18% (w/w) of the phytocannabinoidcontaining fraction.

Non-Cannabinoid Containing Components

The non-cannabinoid components of a phytocannabinoid BDS may play animportant role in the BDS's pharmacology. As such the terpene profile isclassified below. The following tables illustrate the terpene profile ofa CBD chemovar which is representative of a high phytocannabinoidcontaining plant. Five plants were freshly harvested and extracted usingsteam distillation. The principal monoterpene and sesquiterpene arehighlighted in bold.

TABLE 4.5 Monoterpene amount by percentage of total terpene fraction andranges Amount (% of terpene Range Range Range Monoterpenes fraction)(±10%) (±25%) (±50%) Pinene (alpha & 10.56  9.50-11.62  7.92-13.20 5.28-15.84 beta) Myrcene 39.46 35.51-43.41 29.60-49.33 19.73-59.19Limonene 4.14 3.73-4.55 3.11-5.18 2.07-6.21 Beta-ocimene 4.04 3.64-4.443.03-5.05 2.02-6.06 Total 58.20 The monoterpene containing fractioncomprises approximately 52-64% (w/w) of the total terpene fraction.

TABLE 4.6 Monoterpene amount by percentage of monoterpenes Amount (% ofmonoterpene Monoterpenes fraction) Pinene (alpha & beta) 18.14 Myrcene67.80 Limonene 7.12 Beta-ocimene 6.94

The amount of the principal monoterpene myrcene in the monoterpenefraction as a percentage of the monoterpene fraction is approximately61-75% (w/w). The monoterpene fraction also has a secondary monoterpenepinene which is present at approximately 16.3-20% (w/w) of themonoterpene fraction.

TABLE 4.7 Sesquiterpene amount by percentage of total terpene fractionand ranges Amount (% of terpene Range Range Range Sesquiterpenesfraction) (±10%) (±25%) (±50%) Caryophyllenes 29.27 26.34-32.2021.95-36.59 14.64-43.91 (t & oxide) Bergotamene 0.18 0.16-0.20 0.14-0.230.09-0.27 Humulene 7.97 7.17-8.77 5.98-9.96 3.99-11.96 Aromadendrene0.33 0.30-0.36 0.25-0.41 0.17-0.50 Selinene 0.59 0.53-0.65 0.44-0.740.30-0.89 Anon 0.44 0.40-0.48 0.33-0.55 0.22-0.66 Farnesene 1.551.40-1.71 1.16-1.94 0.78-2.33 (Z, E & alpha) alpha Gurjunene 0.120.11-0.13 0.09-0.15 0.06-0.18 Bisabolene 0.39 0.35-0.43 0.29-0.490.20-0.59 Nerolidol 0.43 0.39-0.47 0.32-0.54 0.22-0.65 Diepicedrene-1-0.38 0.34-0.42 0.29-0.48 0.19-0.57 oxide Alpha-Bisabolol 0.16 0.14-0.180.12-0.20 0.08-0.24 Total 41.80 The sesquiterpene containing fractioncomprises approximately 27-32% (w/w) of the total terpene fraction.

TABLE 4.8 Sesquiterpene amount by percentage of sesquiterpenes Amount (%of sesquiterpene Sesquiterpenes fraction) Caryophyllenes (t & oxide)70.02 Bergotamene 0.43 Humulene 19.07 Aromadendrene 0.79 Selinene 1.41Anon 1.05 Farnesene (Z, E & alpha) 3.71 alpha Gurjunene 0.29 Bisabolene0.93 Nerolidol 1.03 Diepicedrene-1-oxide 0.91 Alpha-Bisabolol 0.38

Patent application number PCT/GB2008/001837 describes the production ofa ‘zero cannabinoid’ plant. These plants were produced by selectivebreeding to produce a Cannabis sativa L plant that contained a generallyqualitatively similar terpene profile as a Cannabis sativa L plant thatproduced cannabinoids yet it was devoid of any cannabinoids. Theseplants can be used to produce cannabinoid-free plant extracts which areuseful control plants in experiments and clinical trials. A breakdown ofthe terpene profile produced in the plants can be found in the tablebelow. The primary monoterpenes and sesquiterpene are highlighted inbold.

TABLE 4.9 Monoterpene amount by percentage of total terpene fraction andranges Amount (% of terpene Range Range Range Monoterpenes fraction)(±10%) (±25%) (±50%) Pinene 29.34 26.41-32.27 22.01-36.68 14.67-44.01(alpha & beta) Myrcene 29.26 26.33-32.19 21.95-36.58 14.63-43.89Limonene 5.32 4.79-5.85 3.99-6.65 2.66-7.98 Linalol 4.50 4.05-4.953.38-5.63 2.25-6.75 Verbenol 3.45 3.11-3.80 2.59-4.31 1.73-5.18 (cis &trans) Total 71.87 The monoterpene containing fraction comprisesapproximately 65-79% (w/w) of the total terpene fraction.

TABLE 4.10 Monoterpene amount by percentage of monoterpenes Amount (% ofmonoterpene Monoterpenes fraction) Pinene (alpha & beta) 40.82 Myrcene40.71 Limonene 7.41 Linalol 6.26

The zero cannabinoid plant was found to comprise two principalmonoterpenes; pinene and myrcene. The amount of the principalmonoterpene myrcene in the monoterpene fraction as a percentage of themonoterpene fraction is approximately 37-45% (w/w). The amount of theprincipal monoterpene pinene in the monoterpene fraction as a percentageof the monoterpene fraction is approximately 37-45% (w/w).

Example 7 Use of CBDV (BDS) in the PTZ Model of Generalised Seizures

Methodology as described in Example 2.

CBDV BDS was administered at four doses that yielded a dose of CBDV of50 and 100 mg/kg. Table 7.1 below details the data obtained.

TABLE 7.1 CBDV (mg/kg) Mortality (%) 0 26.3 50 16.7 100 0

As can be seen the CBDV BDS exhibited a trend to decreaseseizure-related mortality.

In contrast to the SAEDs, in all of the experiments using both isolatedCBDV and CBDV BDS, animals did not exhibit any notable side effects.This makes this novel anti-convulsant an attractive compound for useeither alone or in combination in the treatment of epilepsy.

Example 8 Use of THCV (BDS), Isolated THCV and Isolated CBD in Models ofEpilepsy

The data demonstrating the activity of THCV BDS and isolated THCV andCBD are given below in support of the likely benefit of a CBDV extractcontaining CBD and THCV as well as a non-cannabinoid fraction.

General methodology is as described in Example 2.

Results:

The THCV BDS comprised a whole extract of a chemovar in which THCV wasthe predominant cannabinoid. (i.e. it was the major cannabinoid presentin the extract, 80% by weight of the total cannabinoid content). THC wasthe second most prevalent cannabinoid, and was present in significantamounts. (i.e. it comprised greater than 10% by weight of the totalcannabinoid content, being present at about 16%), and there were anumber of minor cannabinoids identified, each comprising less than 2% byweight of the total cannabinoid content as measured by HPLC analysis.The ratio of THCV to THC in this extract is about 5:1.

In fact the THCV content was 67.5% by weight of the extract and the THCcontent was 13.6% by weight of the extract, with the other identifiedcannabinoids in total comprising about 3% by weight of the extract, theremaining 16% comprising non-cannabinoids.

PTZ Pilot Study

Seizures induced by a range of PTZ concentrations (50-100 mg/kg; therange present in the literature) in rats were investigated to determinean optimal dose prior to the investigation of the cannabinoid effect.PTZ doses of:

-   -   50 mg/kg and 60 mg/kg induced very little seizure-like activity        (n=4);    -   70 mg/kg typically induced clonic seizures (score of 3.5; 8 of        13 subjects);    -   80 mg/kg regularly induced tonic-clonic seizures (scores of 4        and 5; 6 of 10 subjects).

Additionally, it was found that repeated dosing with PTZ resulted inincreased sensitivity over time; therefore no experiments were performedon animals that had already received a dose of PTZ.

The effect of THCV BDS on PTZ-induced seizures was first assessedagainst a PTZ dose of 70 mg/kg. As described below, this yielded avehicle control group that did not typically experience severe seizurescores. Therefore THCV BDS was also screened against an 80 mg/kg dose ofPTZ. It was felt that the increased seizure severity experienced byvehicle control animals exposed to 80 mg/kg PTZ was a more appropriatetest of potential anti-convulsant activity.

Effect of THCV BDS on Moderately Severe (70 mg/kg) PTZ-Induced Seizures

Three doses of THCV BDS were assessed against a concentration of PTZknown to induce moderate seizures in rats (70 mg/kg; see pilot, above).The low, medium and high doses of THCV BDS used were 0.37, 3.70 and37.04 mg/kg, and yielded actual THCV doses of 0.25, 2.5 and 25 mg/kgrespectively. These doses were matched by THCV content to those beingused for screening pure THCV against PTZ-induced seizures.

THCV BDS did not have any significant effects on latency to firstmyoclonic jerk or on latency to attaining a severity score of 3.5 on theseizure severity scale (FIG. 14). It should be noted that althoughvalues for both these variables were higher for animals treated withmedium and high dose THCV BDS compared to control, this failed to reachsignificance (P>0.05). Similarly, no significant impact on duration ofseizure was seen (FIG. 15).

The effects of THCV BDS on seizure severity (FIG. 16) and mortality(FIG. 17) in animals that received doses of 70 mg/kg PTZ did not conformto a simple pattern. No animal injected with vehicle-alone exceeded themedian severity score of 3.5 for that group, and no animals died (n=10).

In contrast, 70 mg/kg PTZ induced severe tonic-clonic seizures and deathin 50% of animals injected with a low dose of THCV BDS, demonstrating amedian severity score of 4.75. This increase in severity was notsignificant. However, animals injected with medium and high doses ofTHCV BDS exhibited a lower median severity score and lower mortalityrates than those exposed to low doses (FIGS. 16 & 17). Medium and highdose mortality rates were higher than that of the vehicle group, but notsignificantly so (P>0.05; FIG. 17). However, median severity scores werethe same between medium & high doses (FIG. 16). This pattern of resultssuggested that a further set of experiments, in which THCV BDS wasscreened against a dose of PTZ which would induce severe seizures incontrol (vehicle-treated) animals, was required.

Effect of THCV BDS on Severe (80 mg/kg) PTZ-Induced Seizures

The effects of the same three doses of THCV BDS on seizures induced by80 mg/kg PTZ were assessed. It is worth noting that 80 mg/kg inducedsignificantly more severe seizures than 70 mg/kg in vehicle controlgroups (P=0.009), with median seizure severity scores of 6 and 3.5respectively. THCV BDS did not have a significant effect on latencies toFMJ or a severity score of 3.5 (FIG. 18). Similarly, no effect wasobserved on seizure durations (FIG. 19).

Low dose THCV BDS decreased both seizure severity (FIG. 20) andmortality (FIG. 21) in animals that received doses of 80 mg/kg PTZ.Animals that received low THCV BDS had a lower median severity score(3.5 compared to 6) than vehicle controls. However, this difference wasnot significant (P>0.5). The low THCV BDS dose group also had amortality rate half that of the vehicle control group (30% vs 60%).

Groups treated with medium and high doses of THCV BDS had a lowerseizure severity score of 4.75 (P>0.5 vs control), and a lower mortalityrate of 50%, compared to 6 and 60% respectively.

In vivo Summary and Conclusion

Screening of THCV BDS in the PTZ model did not appear to have anysignificant anti- or pro-convulsant effects on either moderate or severePTZ-induced seizures.

However, a trend towards lower severity and mortality was seen inanimals that received a low dose of THCV BDS prior to induction ofsevere (80 mg/kg PTZ) seizures, compared to vehicle controls.

It is possible that this effect is masked at higher doses of THCV BDS byhigher levels of other cannabinoid constituents (such as THC) present inthe non-THCV content of the THCV BDS. Higher doses of THCV BDS willcontain increasing doses of non-THCV content, such as THC, which mayoppose any potential positive effects of THCV.

Isolated THCV: Effect of Isolated THCV Against PTZ-Induced Seizures

Low (0.025 mg/kg), medium (0.25 mg/kg) and high (2.5 mg/kg) doses ofpure THCV were assessed for their effects on PTZ-induced seizures. It isworth noting at this point, for comparisons to THCV BDS, that differingdoses of pure THCV were used compared to THCV BDS. See Table 8.1 below.

TABLE 8.1 Comparison of THCV BDS and pure THCV doses used in PTZ model“low” dose “medium” dose “high” dose Test CB (mg/kg) (mg/kg) (mg/kg)THCV 0.25 2.5 25 BDS Pure 0.025 0.25 2.5 THCVValues given are for effective THCV content of doses (therefore actualdoses of THCV BDS are approx 1.5 times larger).

80 mg/kg PTZ successfully induced seizures of varying severities inanimals from all 4 experimental groups (n=16 per group). PTZ-inducedseizures led to the death of 44% of animals that received vehicle alone.Groups that received low, medium and high THCV all exhibited lowermortality rates of 41%, 33% and 38% respectively; however these valueswere not significantly different from that of the vehicle group (p>0.05,binomial test).

The mean values for latency to first seizure sign, and to scores of [3]and [5] on the seizure scoring scale used, as well as the duration ofseizure for surviving animals, are described in FIGS. 22A-D.

It can be seen that seizures started later, as shown by increasedlatency to first manifestation of seizure-like behaviour (FIG. 22A) inanimals that received THCV compared to vehicle controls.

The delay of onset was significant at the highest dose of THCV (p=0.02).A similar pattern was seen for latencies to scores of [3] and [5] (FIGS.22B and 22C) with all THCV doses exhibiting increased latencies,reaching a significant level at the highest dose of THCV (p=0.017 and0.013 for [3] and [5] respectively).

It was also observed that duration of PTZ-induced seizures in animalsthat survived the experimental period were significantly shorter afteradministration of the medium dose of THCV compared to vehicle controls(FIG. 22D; p=0.03).

Table 8.2 below displays the values for median seizure severity in eachexperimental group.

TABLE 8.2 Seizure severity and incidence 0.025 mg/kg 0.25 mg/kg 2.5mg/kg Vehicle THCV THCV THCV Median 4.25 3.5 3.5 3.5 severity % noseizure 12.5 5.9 33.3* 18.8

The median maximum severities and % of animals that did not experienceany signs of seizure for each experimental group are given (n=16 foreach value). * indicates significant difference from vehicle group(binomial significance test, P<0.05).

Vehicle control animals exhibited a median seizure severity of 4.25,whereas all groups which received THCV had a median severity score of3.5. This decrease was not significantly different.

12.5% vehicle control animals displayed no indicators of seizure,suggesting these animals did not develop seizures after PTZadministration. A significantly higher number of animals (33.3%)displayed no signs of seizure in the group that received 0.25 mg/kg(Table 5.2; p=0.031). This data suggests that the medium dose of 0.25mg/kg THCV protected against the development of seizures.

In vivo Summary and Conclusion

The effects of the high dose of THCV on latency values suggest that THCVcan delay both onset and seizure development, whilst the significanteffects of the medium dose on the incidence of seizure at medium (0.25mg/kg) THCV doses suggest a significant anticonvulsive action onPTZ-induced seizures.

Isolated CBD

In addition to THCV, CBD was also screened in the PTZ model. The resultsstrongly indicate that CBD (at levels of 100 mg/kg) in this model isanti-convulsant as it significantly decreased the mortality rate andincidence of the most severe seizures compared to vehicle controlanimals.

Effect of Isolated CBD Against PTZ-Induced Seizures

Isolated CBD was injected intra-peritoneally (IP) in the standardvehicle (1:1:18 ethanol: Cremophor: 0.9% w/v NaCI) at doses of 1, 10 and100 mg/kg alongside animals that received vehicle alone at a matchedvolume (n=15 for each group). 60 minutes later PTZ (80 mg/kg, IP) wasadministered.

46.7% of control animals that received vehicle alone died within 30minutes of PTZ administration (FIG. 20). In contrast only 6.7% (only 1of 15) of animals that received 100 mg/kg CBD died, a marked reductionthat proved to be significant (p<0.001).

Additionally only 6.7% of animals that received 100 mg/kg CBDexperienced the most severe seizures (score of 5) in comparison to 53.3%of vehicle control animals, a decrease that was also significant(p<0.001; FIG. 20 in vivo).

In contrast to isolated THCV, no significant increases in latency ofseizure development were observed. However, the marked and significantreductions indicate a striking anti-convulsant effect on PTZ-inducedseizures.

In Vivo Summary and Conclusion

Screening and analysis of isolated CBD in the PTZ model at high dose(100 mg/kg) of CBD on mortality levels and incidence of the most severeseizures suggests that CBD can attenuate the severity of PTZ-inducedseizures.

Overall Conclusion

From the three studies it would appear that both isolated THCV and CBDshow promise as an anti-epileptic for generalized seizure, particularlyclonic/tonic seizure. The data generated for a THCV rich extract,containing other cannabinoids including significant amounts of THC,suggest that the THC may be countering the effect of the THCV and that acannabinoid extract which contains THCV as a major or predominantcannabinoid, but which also contains minimal, or substantially no, THCwould be desirable for treating epilepsy. Furthermore the results withpure CBD suggest that an extract containing significant amounts of bothTHCV and CBD, but again, minimal or substantially no THC may provide anoptimum combination. Accordingly it may prove desirable to prepare aTHCV predominant extract in which THC is selectively, and substantially,removed (to levels of less than a few percent). This could be mixed witha CBD rich extract in which CBD is the major and predominant cannabinoid(also with low levels of THC) to produce an extract with clearlydefined, and significant levels of both THCV and CBD, but withinsignificant levels of THC. Such an extract may contain othercannabinoids and the non-cannabinoid components which result fromextraction, by for example, carbon dioxide as disclosed in WO04/016277,which components may support an “entourage” effect in theendocannabinoid system.

On dosage, a rat/human conversion factor (×6) suggests a CBD daily doseof at least 600 mg (and optionally between 400 mg and 800 mg) and forTHCV at least 1.5 mg (medium) and preferably at least 15 mg (high).

Where a phytocannabinoid extract is to be used, an extract with low ornegligible levels of THC and therapeutically effective levels of THCVand/or CBD is desired.

Example 10 Comparison Between the Anti-Epileptic Activity of IsolatedCBD and CBDV in the Maximal Electroshock Seizure (MES) Model of EpilepsyMethods Preparation of Test and Reference Compounds

The vehicle used in this study was 2:1:17 (ethanol:Cremophor:0.9%^(w)/_(v) NaCI). The test compounds used were cannabidiol (CBD) andcannabidivarin (CBDV). These were made to a solution at the highestconcentration; these were then dissolved in ethanol before combinationwith Cremophor and 0.9% NaCI in the proportion described above. The CBDor CBDV were administered intraperitoneally at a volume of 10 ml/kg bodyweight. The SAED valproic acid (VBA) was dissolved in saline.

Test System

Animal Species/Strain: Mouse/ICR, Microbiological grade: SPF, Supplier:SLC Japan, Inc. Sex: male, Age (at time of testing): 5-7 weeks old,Number of animals: about 5 animals per group. Temperature: 23±2° C.,Humidity: 60±10%, Light conditions: 7 AM to 7 PM for the light period, 7PM to 7 AM for the dark period. Chow and water: Free access to CRF-1(Oriental Yeast Co, Ltd) and tap water.

Experimental Procedures

One day before each experiment, mice were weighed and randomized intoseveral groups in each test. On the morning of the experiment day, bodyweight was measured in order to calculate the administration volume ofeach animal. Vehicle, CBD, CBDV or valproic acid sodium salt wasinterperitoneally administered 30 minutes before electric stimuli.

Maximal electroshock seizures (MES) in mice was induced by a stimulator(UGO BASILE ECT UNIT 7801, Italia) using a current of 30 mA deliveredwith a pulse frequency of 100 Hz for 200 msec through earlap electrodes.The mice were observed for 10 seconds and the incidence of tonichindlimb extension was noted.

Statistical Analysis

All statistical analyses were performed using SAS Software for Windows,Release 9.1 (SAS Institute Japan). The difference of the number(hindlimb extension or deaths) in each group was assessed usingtwo-tailed Fishers exact test. The differences were consideredstatistically significant, when the p value will be less than 0.05.

Results

Almost animals in the vehicle group showed a hindlimb extension inducedby electric stimuli (30 mA for 200 msec). CBD (3-100 mg.kg IP) was notable to inhibit the expression of hindlimb extension with statisticalsignificance. However CBDV (100 and 200 mg/kg IP) significantlyinhibited the expression of hindlimb extension. Meanwhile, the 350 mg/kgof valproic acid blocked the hindlimb extension with statisticalsignificance compared with vehicle group. Tables 10.1 and 10.2 detailthese data.

TABLE 10.1 Effects of CBD and VPA on MES-induced seizure in miceCompound (Dose; mg/kg, i.p.) Incidence of Tonic convulsion Vehicle 5/5CBD (3) 3/5 CBD (10) 4/5 CBD (30) 3/5 CBD (100) 4/5 Valproic acid (350) 0/5** Each group consisted of 5 mice. *= p < 0.05. **= p < 0.01 vsvehicle control (Fisher's exact test)

TABLE 10.2 Effects of CBDV and VPA on MES-induced seizure in miceCompound (Dose; mg/kg, i.p.) Incidence of Tonic convulsion Vehicle 9/10CBDV (50) 9/10 CBDV (100)  3/10* CBDV (200)  3/10* Valproic acid (200)5/10 Valproic acid (350)  1/10** Each group consisted of 10 mice. *= p <0.05. **= p < 0.01 vs vehicle control (Fisher's exact test)

As can be seen from the data above the cannabinoid CBDV unexpectedlydemonstrates greater efficacy as an anti-convulsant in the MES model ofepilepsy than the cannabinoid CBD. Given that the efficacy of CBDV isapproaching that of the SAED valproic acid it is a clear contender foruse an anti-convulsant without producing the side effects that are knownto occur with SAEDs.

Example 11 The Effect of CBDV Upon Motor Function Assessed by LinearlyAccelerating Rotarod Test Methods

Each animal received either CBDV (100 or 200 mg/kg, n=10 for each group)or vehicle (2:1:17 Cremophor:ethanol:saline [n=12] or saline [n=11]) ona given experimental day. Experimental test days were separated by a twoday rest period to allow for clearance of previous compounds. The orderof drug administration was randomised using a standard Latin squaredesign.

60 minutes after CBDV or vehicle administration, animals were placed ona linearly accelerating rotarod (Panlab/Harvard Apparatus, Holliston,USA) that increased in speed from 4 to 40 rpm over a 300 second period.An accelerating protocol was employed to eliminate the need forhabituation to the rotarod, minimising divergence in the resultsobtained for each animal owing to disproportionate improvements inperformance. Each test ended when the animal fell from the rotarod, witheach animal performing three accelerating rotarod runs per experimentalday. Animals were allowed a 5 minute recovery between tests to preventfatigue-induced declines in performance. Mean latency in seconds to fallfrom the rotarod was compared between vehicle control and CBDV groups toassess motor function.

To assess whether there were significant effects on motor functionbetween the two different vehicle treatments, we subjected the data toMann-Whitney U test. Lack of significance in this analysis would allowus to combine vehicle groups thereby reducing duration of testing (i.e.each rat would undertake only one vehicle treatment test day rather thantwo). For analysis of CBDV effects on motor function, data weresubjected to a between-subjects 1-way ANOVA with drug concentration asthe main factor. In all cases, P 0.05 was considered to be significant.

Results

Analysis of Vehicle Treatments: As can be seen in FIG. 24, there was nodifference in the latency to fall between saline and 2:1:17cremaphor:ethanol:saline treated animals (P=0.406). Thus, both vehiclegroups were combined to give us a single vehicle group

Analysis of CBDV effects: As can be seen in FIG. 25, CBDV had no effecton the latency to fall compared to vehicle-dosed animals at any dose(F2, 40=1.421, P=0.253;). Vehicle treated animals remained on therotarod for an average of 111.6 seconds, compared to 86.6 seconds at 100mg/kg CBDV and 110.0 seconds at 200 mg/kg.

Conclusion

These data show that CBDV (100 and 200 mg/kg) had no significant effecton motor control or performance as assessed by accelerating rotarod. Therotarod tests the effect of drugs on the motor behaviour of the rats.Anti-convulsant drugs such as phenobarbital are known to produce adecrease in time that the animals can remain on the rotaroddemonstrating the known side effects of these drugs on motor control.

Thus, the anti-convulsant effects demonstrated in the Examples above, inboth the pentylenetetrazole model of generalised seizures and thepilocarpine model of temporal lobe seizures, are due to thephytocannabinoid CBDV controlling the seizure state without motorside-effects.

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Having thus described several aspects of at least one embodiment of thisinvention, it is to be appreciated various alterations, modifications,and improvements will readily occur to those skilled in the art. Suchalterations, modifications, and improvements are intended to be part ofthis disclosure, and are intended to be within the spirit and scope ofthe invention. Accordingly, the foregoing description and drawings areby way of example only.

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed by the following claims.

All references, including patent documents, disclosed herein areincorporated by reference in their entirety, particularly for thedisclosure referenced herein.

1-19. (canceled)
 20. A formulation comprising CBDV and at least onepharmaceutically acceptable excipient.
 21. A method for the treatment ofepileptic seizures, which comprises administering to a subject in needthereof a therapeutically effective amount of the phytocannabinoid CBDV.22. The method of claim 21, wherein the type of epileptic seizure to betreated is a generalised seizure or a temporal lobe seizure.
 23. Themethod of claim 21, wherein the CBDV is used with one or moretherapeutically effective phytocannabinoids.
 24. The method of claim 23,wherein the one or more therapeutically effective phytocannabinoids isTHCV.
 25. The method of claim 23, wherein the one or moretherapeutically effective phytocannabinoids is CBD.
 26. The method ofclaim 23, wherein the one or more therapeutically effectivephytocannabinoids are THCV and CBD.
 27. The method of claim 21, whereinthe CBDV is in an isolated form.
 28. The method of claim 21, wherein theCBDV is in the form of a botanical drug substance.
 29. The method ofclaim 21, wherein the CBDV is used in combination with a standardanti-epileptic drug.
 30. The method of claim 29, wherein the standardanti-epileptic drug has a mechanism of action which acts via sodium orcalcium channels, or enhances GABAergic inhibition.
 31. The method ofclaim 30, wherein the standard anti-epileptic drug that enhancesGABAergic inhibition is phenobarbital.
 32. The method of claim 30,wherein the standard anti-epileptic drug having a mechanism of actionwhich acts via sodium or calcium channels either: modifies low-thresholdor transient neuronal calcium currents; or reduces high-frequencyneuronal firing and sodium-dependent action potentials and mayadditionally enhance GABA effects.
 33. The method of claim 32, whereinthe standard anti-epileptic drug that modifies low-threshold ortransient neuronal calcium currents is ethosuximide or the standardanti-epileptic drug that reduces high-frequency neuronal firing andsodium-dependent action potentials and may additionally enhance GABAeffects is valproate.
 34. A method for the treatment of epilepticseizures, which comprises administering to a subject in need thereof atherapeutically effective amount of a cannabis plant extract comprisinga phytocannabinoid containing component and a non-phytocannabinoidcontaining component, wherein the phytocannabinoid containing componentcomprises at least 50% (w/w) of the cannabis plant extract and containsas a principal phytocannabinoid, CBDV, and as a secondaryphytocannabinoid, CBD, and wherein the non-phytocannabinoid containingcomponent comprises a monoterpene fraction and a sesquiterpene fraction.35. The method of claim 34, further comprising administering THCV to thesubject.
 36. The method of claim 34, wherein the phytocannabinoidcontaining component comprises 64-78% (w/w) of the cannabis plantextract.
 37. The method of claim 36, wherein the phytocannabinoidcontaining component comprises 52-64% (w/w) CBDV of the totalphytocannabinoid fraction.
 38. The method of claim 37, wherein thephytocannabinoid containing component comprises 22-27% (w/w) CBD of thetotal phytocannabinoid fraction.
 39. The method of claim 37, wherein thephytocannabinoid containing component comprises 3.9-4.7% (w/w) THCV ofthe total phytocannabinoid fraction.